Our long-term goal is to advance knowledge of the neural mechanisms of pain in sickle cell disease (SCD) and develop an effective pharmacologic treatment. The neurobiology of pain in SCD is poorly understood. Several transgenic/knock-out mouse models have been successfully produced and applied to study SCD. However, pain findings were only recently published that indicated increased pain sensitivity in NY1DD mice using a radiant heat tail-flick test. Strikingly, pain in these mice was age-dependent with an onset time around 6 weeks. Unfortunately, the investigators did not report other pain measurement findings or define a pain phenotype in Berkeley sickle transgenic mice, a model of a more severe form of SCD. In our own preliminary studies, we examined Berkeley sickle mice and littermate non-sickle controls in an array of pain tests that are used to study other pain types in animals. In these preliminary studies, we found the presence of tactile allodynia and thermal hyperalgesia and that spinal CaMKII expression and activity were upregulated, similar to what we observed in other mouse models of inflammatory and neuropathic pain. In the latter models, we have identified CaMKIIa to be a critical component leading to persistent pain. We observed that spinal nerve ligation-induced pain behaviors did not develop in CaMKIIa mutant mice. Based on these encouraging data, we propose to extensively characterize pain behaviors in Berkeley mice by employing standard pain tests for spontaneous pain behaviors and those evoked by thermal or mechanical stimuli (von Frey, Hargreaves, hot-plate, tail-flick, cold allodynia) and inflammatory stimuli (formalin, complete Freund's adjuvant) from shortly after birth through adulthood or when responses plateau. Some of these pain tests are used in our ongoing human studies of SC pain using quantitative sensory testing (QST). Intentionally, these similarities will allow us to interpret findings from the SCD transgenic mouse model with consideration of findings from other mouse models of pain and human SCD pain. We will use real time PCR, immunoblotting, immunohisto-chemistry, and enzymatic kinetics methods to systematically examine the expression and activity of CaMKIIa and total CaMKII in sickle and control mice and correlate the changes in this potential biomarker with the onset of pain. To directly test the hypothesis that spinal CaMKIIa is a molecular mechanism that promotes and maintains the manifestation of chronic pain in SCD, we will conduct pharmacological studies to inhibit CaMKIIa using chemical, small interfering RNA (siRNA), and gene knockout methods. We propose to test in these pharmacological studies a clinically used orally available drug that we have found to be a CaMKII inhibitor and to reduce pain behaviors in inflammatory and neuropathic pain models. Our secondary strategy is to conduct a pilot translational study to identify safety issues and clinical potential of this CaMKII inhibitor by characterizing sensory pain in humans with quantitative sensory testing and a computerized self-report tool. The significance of this proposal is that it may ultimately lead to pharmacological interventions that target the CaMKII-pathway. (End of Abstract)